Challenge 23: Retinal 3D Retinal 3D: A Physiologically-Competent - - PowerPoint PPT Presentation

Challenge 23: Retinal 3D

Retinal 3D:

A Physiologically-Competent Human Retinal 3D Model

Launch Meeting 08 September 2016 Stefan Kustermann (Roche), Phil Hewitt (Merck), Francois Pognan (Novartis)

• r Marianne Uteng (Novartis) (who will be present at the meeting)

The Challenge

Establish a human 3D retinal cell model:

• Physiologically-competent and predictive of human physiology
• Consist of all major cell types of the retina : Müller- and micro-glia, RPE

and neurons (including photoreceptors)

• Enable cellular interplay
• Recapitulates key morphological and functional features
• Provide a panel of relevant readouts for functional testing

Why was this Challenge developed? Scientific – Business - 3Rs drivers

• Over 60 million people worldwide are blind
• Leading causes of blindness in the industrial world is AMD
• Globally, in ophthalmology, more than 600 R&D projects running – with more to

come

• Large scientific and business impact of a novel human 3D retina model
• Current in vitro models have major limitations:
• cells are immature, no interplay between different cell types, functionality

limited, lack human relevance etc.

• strong need for better models of human relevance
• majority of studies for efficacy and safety testing in ophthalmological drug

development are performed in animals due to lack of relevant in vitro models

• Ocular safety studies in vivo use up to 20 animals per compound
• strong impact on 3Rs if human relevant model can be applied for R&D

Current state of the art

• Single cells:
• 2D cell line models (human and rodent): ARPE19, Mio-M1, 661W

etc.

• Complex models:
• Retinal explants of mouse, rat and pig
• Stem cell derived models:
• retina 3D spheroids and isolated cells thereof
•  Lack of a human relevant, complex retinal 3D

model

Established complex retinal cell models: Retinal explant cultures

• remains viable as an explant in

serum-supplemented growth media for more than 4 weeks

• histotypic development of neonatal

as well as preservation of late postnatal mouse retinal structure during long-term culture

• also feasible with:
• Rat (Pinzon-Duarte et al 2000)
• Pig (Wang et al 2011)
• Zebrafish (Kustermann et al 2008)
• Not feasible with human retina

due to lack of tissue

e.g. Caffé et al. 2001

Established complex retinal cell models: Whole embryonic-body culture

• Mouse ES cells generate fully

stratified neuro-retinas after 24 days

• Photoreceptors can be isolated

and transplanted into eyes in vivo and morphologically integrate

• Phenotype of cells is pre-

mature and long differentiation time for e.g. photoreceptors and glia cells (humans ~6 month)

e.g. Eiraku et al. 2011; Gonzalez-Cordero et al. 2013

Deliverables Phase 1

Mimic ic the e morphol rphology

• gy and

d physio siology

• gy of the

e mature ure huma man n retin tina:

• Morphological resemblance of layered structure of the retina

i.e. plexiform and nuclear layers

• Sustained expression of cell specific (mature) markers by IHC
• f all implemented cell types
• Basic functional characterization: e.g. synapse formation

between neurons by IHC, tight junction expression between RPE cells, phagocytotic activity of microglial cells

What we don’t want

• A model that requires complex and time consuming set-up,

impeding parallel testing of multiple conditions

• Incorporate only cytotoxicity as a readout
• Incorporate material which is known to show strong compound

adsorption (e.g. PDMS)

• Require specific legal work for acquisition of source cells and materials

(i.e. individual informed consent for each study/experiment)

Deliverables Phase 2 “Must have”

• Thorough functional and morphological characterisation of the

retinal model including recapitulation of drug induced retinal toxicities:

• Development of additional methods to address function and/or phenotypic

changes of the different cell types in culture

• Provision of accessible morphological and functional readouts and be compatible

with standard microscopes

• Recapitulation of retinal toxicities of known drugs (e.g. Chloroquine, …)
• Amenity to the testing of compounds in parallel (multi-well-plate setting)
• Easy to implement in an industry laboratory setting, reproducible and easily

transferrable between laboratories

• Assessment of inter/intra-laboratory reproducibility
• Adaptability to other species relevant for safety assessment (rodent and /or non-

rodent)

• Evidence of cost efficiency and applicability to individual and longer term

experiments

Deliverables Phase 2 “nice to have”

• Demonstration of the barrier function of outer and/or inner-limiting membrane
• A functional blood-retina barrier
• Vasculature (endothelial cells) to mimic neo-vascularization and leakage of blood

vessels to model disease phenotypes such as Diabetic Macular Edema

• Clear strategy how to commercialise the results into a product or service

Sponsors’ contributions

• Expertise in ophthalmology and in vitro models including specifications

for an in vitro model which is fit for purpose for drug testing in an industry setting

• Compounds and knowledge of compounds where available for evaluation
• f both the pharmacological and toxicological performance of the in vitro

retinal test system

• Potential for in-house testing using the system to test transferability and

reproducibility of the 3D retinal in vitro model

The Sponsors are happy to discuss the challenge and potential applications with people in the run up to the submission deadline Sponsor contacts are: Roche Dr Stefan Kusterman stefan.kustermann@roche.com Merck Dr Philip Hewitt Philip.Hewitt@merckgroup.com Novartis Dr Francois Pognan francois.pognan@novartis.com Dr Marianne Uteng marianne.uteng@novartis.com Bayer Dr Thomas Steger-Hartmann thomas.steger-hartmann@bayer.com NC3Rs crackitenquiries@nc3rs.org.uk

Thank You